We previously demonstrated that age-, disease- and therapy-associated effects on thymic function are fundamental causes of diminished T cell immune reconstitution following lymphocyte depletion. Thus, one can overcome this limitation by either enhancing thymic function or enhancing the efficiency of thymic-independent pathways of immune reconstitution. We have focused our efforts on clinical development of recombinant human IL-7, which is a potent immunorestorative that works predominantly by enhancing thymic-independent immune reconstitution.In FY11 we published the clinical results of our first clinical trial of rhIL7 in humans, which demonstrated potent effects on early B cells and documented prolonged T1/2 of this cytokine agent. These results were surprising, as previous work had demonstrated cytokines to be very short lived in vivo (Sportes et al, Clin Can Res 2010). The results also clearly documented the potent immunorestorative effects of rhIL7 and demonstrated minimal toxicity, thus setting the stage for use of this agent in a variety of settings aimed at enhancing immune reconstitution, vaccine responses and antiviral responses in the setting of chronic viral infection. Based upon the results of this study, over 20 studies worldwide are underway utilizing IL7 as an immunomodulator and this agent holds great promise for eventual approval for use in a variety of clinical settings. In FY11 we also published a high impact summary article detailing the promising of interleukin-7 for clinical use in the premier immunology review journal worldwide (Mackall et al, Nat Rev Immunology).The first major accomplishment of this project during FY12 was completion of studies demonstrating an important role for soluble IL7 receptor in modulating bioactivity of IL7. We initiated a project studying the biology of IL7 receptor because polymorphisms in this gene have been linked to differential susceptibility to multiple sclerosis. Despite this clear genetic evidence of a role for IL7R in modulating susceptibility to disease, there was essentially no understanding of the biology responsible for these findings. Through these studies, we found that the genetic variation in IL7R that predisposes to autoimmunity results in a higher splicing rate of this molecule, with increased copies of the mRNA that is lacking the transmembrane domain (e.g. exon 6). This leads to the prediction that the polymorphisms predisposing to autoimmunity would be associated with higher levels of soluble IL7R. Indeed, using cohorts of healthy and MS-afflicted patients, we found that autoimmunity predisposing polymorphisms leads to higher circulating levels of soluble IL7Ra. Furthermore, when soluble IL7Ra is present, IL7 induces diminished short-term high burst signaling, and increased IL-7 signaling over the longer term (e.g. days). This is due to diminished IL7 consumption in the presence of soluble IL7Ra. We also observed qualitative differences in the downstream events induced by rhIL7 in the presence or absence of soluble receptor, such that diminished levels of negative regulators were induced. Together, this led to the hypothesis that polymorphisms in IL7R increase the risk of autoimmunity of enhancing the bioactivity of IL-7, in part by diminishing consumption and increasing availability. Using clinical samples from healthy donors and from patient with multiple sclerosis, we observed that soluble IL7R levels are tightly regulated by genetic polymorphisms in IL7Ra and that in patients with multiple sclerosis, IL-7 levels themselves are also modulated by polymorphisms in IL7Ra. Finally, we observed in a mouse model of autoimmune encephalitis, that co-administration of sIL7Ra significantly increases the potency of IL-7 induced disease, which provided a clear basis for implicating soluble IL7Ra levels in susceptibility to multiple sclerosis. These studies provide fundamental insights into the important role that IL7R plays in modulating IL7 bioactivity in vivo, elucidate a clear basis to explain how genetic polymorphisms increase susceptibility to multiple sclerosis when increased levels of soluble IL7R are present and lead to the prediction that co-treatment with IL7 plus sIL7Ra will enhance the potency of IL7 as a therapeutic agent. Finally, these data reveal that under normal circumstances, humans have a profound molar excess of soluble IL7Ra to IL7 itself. This is fundamentally different that the relationship between soluble receptor found for IL2 and IL15 and likely provided the explanation for the prolonged half-life of IL7 when used as a therapeutic agent.The second major accomplishment of this project in FY12 was administration of rhIL7 to a large cohort of children with cancer as part of a clinical trial of immune restoration after primary front line therapy. This represents the only clinical experience with rhIL7 in children thus far. We demonstrated the agent to be safe and highly effective at increasing the pace and degree of immune reconstitution following intensive chemotherapy for cancer. Furthermore, as discussed in Project III, exciting clinical results suggest that this approach may improve survival in this high-risk population.